TLC analyses of the hydrolyzed products of AmyM action. (A) Hydrolysis activity of AmyM. Each substrate (0.5%, wt/vol) was incubated without (− lanes; substrate controls) or with (+ lanes) the enzyme (2 U/ml) at 42°C for 12 h. G1, glucose; G2, maltose; G3, maltotriose; G4, maltotetraose; G5, maltopentaose; α-CD, α-cyclodextrin; β-CD, β-cyclodextrin; γ-CD, γ-cyclodextrin; SS, soluble starch; Pul, pullulan. Lane Std, maltooligosaccharide standards from glucose (G1) to maltopentaose (G5); lane P, panose standard. (B) Time course analysis of hydrolyzed products of soluble starch. Soluble starch (1%, wt/vol) was incubated with the enzyme (0.2 U/ml) at 42°C. The spots larger than maltotriose generated at later stages were confirmed by HPIC as branched oligosaccharides. (C) Simplified diagram describing the hydrolysis of starch, β-cyclodextrin, and pullulan by AmyM. Circle, glucose unit; slashed circle, reducing end of the molecule; dotted arrows, cutting sites by AmyM; line, α-(1,4) glycosidic linkage; ↓, α-(1,6) glycosidic linkage. n, arbitrary natural number.